The overall goal of this grant proposal is to discover novel anticancer drugs based on designing small synthetic molecules that inhibit STAT3 dimerization and aberrant function in tumors that depend on STAT3 for survival. Overwhelming pre-clinical evidence points to the pivotal involvement of STAT3 in promoting several hallmarks of cancer including uncontrolled cell proliferation, apoptosis evasion, angiogenesis, and metastasis. Furthermore, clinically, in patient tumors, STAT3 is found persistently hyper- activated (tyrosine phosphorylated) and this is associated with resistance to chemotherapy and poor prognosis. In tumor cells which harbor aberrantly activated STAT3, inhibiting STAT3 function with dominant negative STAT3 beta, knocking down its expression or inhibiting its activation by STAT3 activation inhibitors results in reversal of malignant transformation and/or induction of apoptosis. STAT3 tyrosine phosphorylation and subsequent dimerization are required for STAT3 activation. The hypothesis upon which this grant is based is that STAT3 dimerization selective inhibitors will suppress STAT3 aberrant function and inhibit STAT3-mediated up-regulation of anti-apoptotic and/or pro-oncogenic proteins, and will suppress tumor growth and induce apoptosis. To test this hypothesis the following specific aims are proposed: 1) To use structure-based Virtual HTS and molecular modeling and to perform experimental HTS to identify and design potent and selective STAT3 dimerization inhibitors. 2) To determine the ability of leads from Specific Aim 1 to potently and selectively inhibit STAT3 dimerization, STAT3 tyrosine phosphorylation, STAT3-DNA-binding and STAT3-dependent transcriptional activities in whole cells, 3) To determine if the highly selective STAT3 dimerization inhibitors identified in Specific Aim 2 are able to inhibit potently and selectively STAT3- dependent oncogenic signaling and suppress malignant transformation in a STAT3-dependent manner in human cancer cells, and 4) To evaluate the anti-tumor activity, pharmacodynamics, pharmacokinetics and toxicity in animal models of potent and selective STAT3 dimerization inhibitors. The studies proposed will provide chemical probes that will allow us to enhance our understanding of the importance of STAT3 in human oncogenesis, and ultimately, the studies will lead to the discovery of STAT3 inhibitors with pro-apoptotic and anti-tumor activities, and will broaden the spectrum of human tumors that can be successfully treated.